We investigate T cell development and function. Most T cells recognize peptide antigens presented by class I (MHC-I) or class II (MHC-II) classical Major Histocompatibility Complex molecules, through a surface antigen receptor (TCR). This TCR comprises two antigen-specific chains, &#945; and &#946;, whose coding sequence is generated during early thymocyte development by the random rearrangement of TCR&#946; and TCR&#945; loci. TCR antigen recognition is aided by two surface glycoproteins called coreceptors: CD4, which binds MHC-II, or CD8, which binds MHC-I. Coreceptor expression on mature T cells is mutually exclusive and strongly correlates with both MHC specificity and functional (helper vs. cytotoxic) differentiation. That is, the general rule is that MHC I-specific T cells are CD4CD8+ and cytotoxic, whereas MHC II-specific T cells are CD4+CD8 and helper. This double correspondence is essential to the proper function of the immune system; understanding how it is established during T cell development in the thymus and maintained in mature T cells are key questions of developmental T cell biology. <br>CD4 and CD8 T cells differentiate in the thymus from precursors expressing both CD4 and CD8 molecules (double positive, DP). The differentiation of DP thymocytes into mature T cells has been the main focus of our research. Our current approach to this question began with large-scale gene expression analyses performed a few years ago, in which we compared gene expression in thymocytes undergoing selection to that in pre-selection DP cells. These analyses led us to identify a zinc finger DNA-binding protein, previously known as cKrox or Thpok and now referred to as Zbtb7b, as a major CD4-differentiating factor. We showed that the Zbtb7b gene is expressed in a CD4-specific manner in the T cell compartment (although its expression is not restricted to lymphocytes) and that it is up-regulated during the positive selection of CD4 but not of CD8 T cells. We also showed that transgenic expression of Zbtb7b during T cell development forces selected thymocytes into the CD4 lineage, even if they are MHC I-specific and thus would normally differentiate into CD8 cells. These results (Sun et al., Nat Immunol, 2005), and similar findings reported by others, identify Zbtb7b as a major CD4-differentiating factor and form the basis for our current research plans. <br> We are pursuing these investigations along three main lines. First, we have generated a number of mouse models to further study the role of Zbtb7b in development. This includes Zbtb7b-deficient mice and lines transgenic for a GFP-based bacterial artificial chromosome reporter for Zbtb7b expression. We are currently investigating lymphocyte development in these animals. <br> Second, we further examined Zbtb7bs ability to repress the CD8 differentiation program. To this end, we introduced this protein by retroviral transduction into mature (peripheral) CD8 T cells in which it is normally not expressed. These analyses indicate that Zbtb7b represses the expression of CD8 and of characteristic cytotoxic genes, including perforin and granzyme B. Conversely, we found a noticeable induction of helper-specific genes (including those encoding IL-2 and IL-4) in Zbtb7b-transduced CD8 T cells. These observations (Jenkinson et al., J Exp Med, 2007) identify Zbtb7b as a potent repressor of the CD8-cytotoxic gene expression program. Furthermore, the fact that Zbtb7b repression of this program is still active in post thymic CD8 cells suggests that linage differentiation is not entirely locked in post-thymic T cells by epigenetic mechanisms but rather remains, at least in part, sensitive to some of the transcription factors that decide lineage choice in the thymus. <br> Third, we are investigating molecular aspects of Zbtb7b-directed CD4 differentiation. We recently found that Zbtb7b antagonizes the repression of CD4 expression by the transcription factor Runx3, an event characteristic of CD8 differentiation, and we are currently exploring the mechanisms of this effect